Natural hybridization frequently occurs in plants and can facilitate gene flow between species, possibly resulting in species refusion. However, various reproductive barriers block the formation of hybrids and maintain species integrity. Here, we conducted a field survey to examine natural hybridization and reproductive isolation (RI) between sympatric populations of Primula secundiflora and P. poissonii using ten nuclear simple sequence repeat (SSR) loci. Although introgressive hybridization occurred, species boundaries between P. secundiflora and P. poissonii were maintained through nearly complete reproductive isolation. These interfertile species provide an excellent model for studying the RI mechanisms and evolutionary forces that maintain species boundaries.